The invention relates to a process for the fermentative preparation of L-amino acids, in particular L-threonine, using coryneform bacteria in which the glyA gene is attenuated. All references cited herein are expressly incorporated by reference. Incorporation by reference is also designated by the term xe2x80x9cI.B.R.xe2x80x9d following a citation.
L-Amino acids are used in animal nutrition, in human medicine and in the pharmaceuticals industry. It is known that amino acids are prepared by fermentation from strains of coryneform bacteria, in particular Corynebacterium glutamicum. Because of their great importance, work is constantly being undertaken to improve the preparation processes.
Improvements to the processes can relate to fermentation measures, such as, for example, stirring and supply of oxygen, or the composition of the nutrient media, such as the sugar concentration during the fermentation, or the working up to the product form by, for example, ion exchange chromatography, or the intrinsic output properties of the microorganism itself.
Methods of mutagenesis, selection and mutant selection are used to improve the output properties of these microorganisms. Strains which are resistant to antimetabolites, such as the threonine analogue xcex1-amino-xcex2-hydroxyvaleric acid (AHV), or are auxotrophic for metabolites of regulatory importance and produce L-amino acids such as threonine are obtained in this manner.
Recombinant DNA techniques have also been employed for some years for improving the strain of Corynebacterium glutamicum strains which produce L-amino acids, by amplifying individual amino acid biosynthesis genes and investigating the effect on the L-amino acid production. Review articles in this context are to be found, inter alia, in Kinoshita (Glutamic Acid Bacteria, in: Biology of Industrial Microorganisms, Demain and Solomon (Eds.), Benjamin Cummings, London, UK, 1985, 115-142) I.B.R., Hilliger (BioTec 2, 40-44 (1991)) I.B.R., Eggeling (Amino Acids 6, 261-272 (1994)) I.B.R., Jetten and Sinskey (Critical Reviews in Biotechnology 15, 73-103 (1995)) I.B.R. and Sahm et al. (Annuals of the New York Academy of Science 782, 25-39 (1996)) I.B.R.
An object of this invention is to provide new and improved processes for the fermentative preparation of L-amino acids with coryneform bacteria. L-Amino acids are used in human medicine and in the pharmaceuticals industry, in the foodstuffs industry and especially in animal nutrition. There is therefore a general interest in providing new improved processes for the preparation of amino acids. Where L-amino acid is mentioned below, this means L-threonine or L-isoleucine.
The invention provides a process for the fermentative preparation of L-amino acids using coryneform bacteria in which at least the nucleotide sequence which codes for the glyA gene product (glyA gene) is attenuated, and in particular expressed at a low level. The desired product is concentrated in the medium or in the cells and the L-amino acid is isolated. The strains employed preferably already produce L-amino acids before attenuation of the glyA gene.